Epigenetic and Transcriptional Rewiring of Circadian Clock Genes by Polycyclic Aromatic Hydrocarbons: Implications for Lung and Breast Cancer Initiation and Progression

1. Introduction

Environmental exposures are increasingly recognized as critical determinants of human health, particularly in the context of chronic diseases such as cancer [1]. Among environmental pollutants, polycyclic aromatic hydrocarbons (PAHs) have attracted significant attention due to their widespread distribution and well-established carcinogenic potential [2]. At the same time, the circadian rhythm, an endogenous timekeeping system that orchestrates physiological processes over a 24-hour cycle, has emerged as a key regulator of cellular homeostasis. Disruption of circadian timing, often referred to as chronodisruption, has been implicated in the pathogenesis of multiple diseases, including metabolic disorders and cancer [3]. The integration of these two domains has led to the emergence of environmental chronotoxicology, a field that examines how environmental toxicants interact with biological timing systems to influence disease risk and progression [4,5].

Recent advances suggest that PAHs may exert part of their carcinogenic effects by interfering with circadian clock machinery through molecular pathways involving the aryl hydrocarbon receptor (AhR), oxidative stress signaling, and epigenetic modifications. These interactions can disrupt rhythmic gene expression, thereby altering key cellular processes such as DNA repair, cell cycle regulation, and apoptosis, which are tightly controlled by the circadian system [6]. Understanding this interplay is essential for elucidating the mechanisms linking environmental exposures to cancer initiation and progression.

1.1. Overview of Polycyclic Aromatic Hydrocarbons Exposure

Polycyclic aromatic hydrocarbons (PAHs) are a diverse class of organic compounds composed of multiple fused aromatic rings, primarily formed during the incomplete combustion of organic materials [7]. These compounds are ubiquitous in the environment, originating from both natural and anthropogenic sources. Industrial activities such as petrochemical processing, aluminum production, and waste incineration represent major contributors to environmental PAH contamination. In addition, combustion-related processes, including vehicle emissions, fossil fuel burning, and biomass combustion, significantly elevate atmospheric PAH levels, particularly in urban settings. Tobacco smoke also constitutes a major indoor source of PAHs, while dietary intake through consumption of grilled or smoked foods represents another important exposure pathway [8,9].

Human exposure to PAHs occurs predominantly through inhalation, ingestion, and dermal contact. Inhalation is considered the primary route, especially in polluted environments where PAHs are bound to airborne particulate matter [10,11] . Ingestion of contaminated food and water contributes substantially to total exposure, particularly in populations with high consumption of charred foods. Dermal exposure is also relevant in occupational settings, such as in workers handling asphalt, coal tar, or industrial by-products (Figure 1) [12,13]. Following entry into the body, PAHs are readily absorbed due to their lipophilic nature and distributed to various tissues, with a tendency to accumulate in lipid-rich compartments. They undergo metabolic activation primarily via cytochrome P450 enzymes, especially CYP1A1 and CYP1B1, resulting in the formation of reactive intermediates capable of forming DNA adducts and inducing mutagenesis. This metabolic activation is a key step in PAH-induced carcinogenesis (Table 1) [14].

1.2. The Biology of the Circadian Rhythm

The circadian rhythm is an intrinsic timekeeping system that enables organisms to anticipate and adapt to daily environmental changes [17]. This approximately 24-hour cycle regulates a wide range of physiological processes, including sleep–wake patterns, hormone secretion, metabolism, immune responses, and cellular repair mechanisms. In mammals, the circadian system is hierarchically organized, with a central pacemaker located in the suprachiasmatic nucleus (SCN) of the hypothalamus. The SCN synchronizes peripheral clocks present in nearly all tissues, including the liver, lung, and breast, thereby ensuring coordinated regulation of tissue-specific functions. External environmental cues, known as zeitgebers, such as light, feeding cycles, and temperature, play a crucial role in entraining the circadian system to the external environment [18,19].

At the molecular level, circadian rhythms are governed by interconnected transcriptional–translational feedback loops involving a set of core clock genes and proteins [19,20]. The transcription factors CLOCK and BMAL1 form heterodimers that drive the expression of target genes, including the period (PER1–3) and cryptochrome (CRY1–2) genes. The PER and CRY proteins accumulate in the cytoplasm, translocate back to the nucleus, and inhibit CLOCK–BMAL1 activity, thereby forming a negative feedback loop (Figure 2). Additional regulatory loops involving nuclear receptors such as REV-ERB and ROR further stabilize and fine-tune circadian oscillations. These molecular oscillations generate rhythmic gene expression patterns that regulate downstream biological processes essential for maintaining cellular homeostasis. Disruption of these tightly regulated feedback mechanisms can lead to profound physiological consequences, including increased susceptibility to disease (Table 2) [21,22].

1.3. Concept of Chronotoxicity and Cancer Linkage

Chronotoxicity refers to the time-dependent variation in the toxicity of chemical agents, reflecting the influence of circadian rhythms on the absorption, distribution, metabolism, and excretion of xenobiotics. A closely related concept, chronodisruption, describes the disturbance of normal circadian organization due to environmental, behavioral, or physiological factors. Such disruptions can manifest as altered sleep–wake cycles, hormonal imbalances, and loss of rhythmic gene expression, ultimately leading to systemic physiological dysregulation [25].

Emerging evidence indicates that environmental toxicants, including PAHs, can function as chronotoxic agents by interfering with circadian clock function at multiple levels [26,27]. Mechanistically, PAHs activate the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor that shares structural and functional similarities with circadian regulators such as CLOCK and BMAL1. This interaction enables competitive or cooperative binding to DNA regulatory elements, thereby altering circadian gene expression. In addition, PAHs induce oxidative stress and inflammatory signaling pathways, including reactive oxygen species (ROS) production and NF-κB activation, which can further disrupt redox-sensitive components of the circadian machinery. Epigenetic modifications, such as DNA methylation and histone remodeling, also contribute to sustained alterations in circadian gene expression following toxicant exposure [28,29,30,31].

The link between circadian disruption and cancer is supported by both experimental and epidemiological evidence. Circadian clock genes regulate critical cellular processes, including DNA damage response, cell cycle progression, and apoptosis [21,32]. Disruption of these regulatory mechanisms can lead to genomic instability, uncontrolled cell proliferation, and impaired programmed cell death, all of which are hallmarks of cancer [33]. Epidemiological studies, particularly those involving shift workers, have demonstrated an increased risk of cancers such as lung and breast cancer, leading to the classification of circadian disruption as a probable carcinogen by the International Agency for Research on Cancer [4,34,35,36,37,38,39,40]. In this context, PAH-induced chronodisruption represents a mechanistic bridge linking environmental exposure to cancer initiation and progression, highlighting the importance of integrating temporal biology into toxicological and carcinogenic risk assessment.

2. Molecular Mechanisms of PAH-Induced Circadian Disruption

The ability of polycyclic aromatic hydrocarbons (PAHs) to perturb circadian homeostasis is mediated through a complex network of molecular pathways that integrate xenobiotic sensing, transcriptional regulation, redox signaling, and inflammatory responses. Central to this process is the activation of the aryl hydrocarbon receptor (AhR), a ligand-activated transcription factor that serves as a key mediator of environmental chemical signaling. The convergence of AhR signaling with circadian clock machinery, alongside oxidative stress and inflammatory cascades, creates a multifaceted mechanism through which PAHs disrupt circadian rhythmicity at both molecular and physiological levels [6,26,41,42].

2.1. Activation of Aryl Hydrocarbon Receptor (AhR) Signaling

The aryl hydrocarbon receptor (AhR) is a cytosolic transcription factor that plays a central role in the detection and metabolic processing of environmental toxicants, including PAHs. In its inactive state, AhR resides in the cytoplasm in association with chaperone proteins such as heat shock protein 90 (HSP90), XAP2, and p23 [43]. Upon exposure to PAHs, lipophilic ligands readily diffuse across the cell membrane and bind to AhR, inducing a conformational change that facilitates its dissociation from the chaperone complex. This ligand-bound AhR then translocates into the nucleus, where it dimerizes with the aryl hydrocarbon receptor nuclear translocator (ARNT) [14,44].

Once in the nucleus, the AhR–ARNT complex binds to specific DNA sequences known as xenobiotic response elements (XREs), initiating the transcription of a battery of target genes involved in xenobiotic metabolism, including cytochrome P450 enzymes such as CYP1A1, CYP1A2, and CYP1B1 [43]. While this pathway is essential for detoxification, it also leads to the metabolic activation of PAHs into reactive intermediates capable of inducing DNA damage. Importantly, emerging evidence suggests that AhR signaling extends beyond xenobiotic metabolism to influence broader transcriptional networks, including those governing circadian regulation. The structural similarity between AhR and core circadian transcription factors enables functional overlap and potential interference with circadian gene expression programs [30,45,46,47].

2.2. Crosstalk Between AhR and Circadian Clock Machinery

A critical mechanism underlying PAH-induced circadian disruption is the molecular crosstalk between AhR signaling and the circadian clock system. Both AhR and core circadian proteins such as CLOCK and BMAL1 belong to the basic helix–loop–helix PER-ARNT-SIM (bHLH-PAS) family of transcription factors, which allows them to share common dimerization partners and DNA-binding motifs. This structural homology provides a basis for direct interaction and competition between these pathways [6,48,49].

Experimental studies have demonstrated that activation of AhR can interfere with the formation and activity of the CLOCK–BMAL1 heterodimer, the central transcriptional activator of circadian gene expression. AhR may compete with CLOCK or BMAL1 for binding to ARNT, thereby limiting the availability of this cofactor for circadian transcriptional complexes. In addition, AhR activation has been shown to repress the transcription of key circadian genes such as PER1, PER2, and BMAL1, either directly through promoter interactions or indirectly via downstream signaling pathways [6,30,41].

Conversely, circadian clock components can also regulate AhR signaling, indicating a bidirectional relationship [6]. CLOCK–BMAL1 complexes may modulate the rhythmic expression of AhR target genes, thereby influencing the temporal dynamics of xenobiotic metabolism. This reciprocal regulation underscores the integration of environmental sensing with circadian timing systems. However, sustained activation of AhR by persistent environmental pollutants such as PAHs can overwhelm this balance, leading to prolonged disruption of circadian transcriptional networks and loss of rhythmic gene expression (Figure 3) [30,41,50].

2.3. Oxidative Stress and Inflammatory Signaling

In addition to direct transcriptional interference, PAHs contribute to circadian disruption through the induction of oxidative stress and inflammatory signaling pathways. The metabolic activation of PAHs via cytochrome P450 enzymes generates reactive oxygen species (ROS), leading to a state of redox imbalance within the cell. Elevated ROS levels can damage cellular macromolecules, including DNA, lipids, and proteins, and also act as signaling molecules that modulate gene expression [28,51,52,53]. Circadian clock components are highly sensitive to cellular redox status, as several clock proteins contain redox-responsive domains that influence their stability and activity [54,55]. Oxidative stress can therefore alter the function of key circadian regulators such as CLOCK, BMAL1, and PER proteins, resulting in disrupted oscillatory patterns. Furthermore, ROS-mediated activation of signaling pathways such as nuclear factor kappa B (NF-κB) plays a significant role in linking inflammation to circadian dysregulation. NF-κB is a transcription factor that regulates the expression of pro-inflammatory cytokines and has been shown to interact with circadian clock genes, either by directly binding to their promoters or by modulating their transcriptional activity [56,57,58,59].

Chronic activation of inflammatory pathways further exacerbates circadian disruption by creating a feedback loop in which inflammation and clock dysfunction reinforce each other. Pro-inflammatory cytokines such as tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) can suppress the expression of core clock genes, leading to a sustained loss of circadian rhythmicity [60,61]. This interplay between oxidative stress, inflammation, and circadian regulation represents a key mechanism through which PAHs exert their chronotoxic effects (Figure 4).

2.4. Temporal Disruption and Loss of Circadian Oscillation

The cumulative effects of AhR activation, transcriptional interference, oxidative stress, and inflammation ultimately manifest as alterations in the temporal organization of circadian rhythms [6]. One of the hallmark features of PAH-induced circadian disruption is phase shifting, in which the timing of peak gene expression is advanced or delayed relative to the normal circadian cycle. Such phase shifts can desynchronize internal biological clocks from external environmental cues, leading to physiological misalignment [62,63,64]

Another critical consequence is the reduction in amplitude of circadian oscillations, characterized by diminished fluctuations in gene expression levels over the 24-hour cycle [26,28]. Reduced amplitude reflects a weakening of the underlying transcriptional feedback loops and is often associated with impaired physiological function. In more severe cases, sustained exposure to PAHs can lead to arrhythmicity, a complete loss of circadian oscillations, resulting in the breakdown of temporal regulation across multiple biological systems [5].

At the cellular level, these temporal disruptions have profound implications for processes such as DNA repair, cell cycle progression, and metabolic regulation, all of which are under circadian control. The loss of coordinated timing can therefore create a permissive environment for genomic instability and tumorigenesis (Table 3). Importantly, the extent of circadian disruption may depend on factors such as the duration and timing of exposure, highlighting the importance of temporal dynamics in environmental toxicology [33,65].

3. Epigenetic and Transcriptional Rewiring of Circadian Clock Genes

The disruption of circadian rhythms by polycyclic aromatic hydrocarbons (PAHs) extends beyond transient signaling interference to encompass stable and heritable changes in gene expression mediated through epigenetic and transcriptional mechanisms [26]. Epigenetic regulation including DNA methylation, histone modifications, and non-coding RNA activity plays a central role in maintaining the rhythmic expression of circadian clock genes [66]. However, environmental toxicants such as PAHs can alter these regulatory layers, leading to persistent reprogramming of circadian transcriptional networks. This epigenetic rewiring not only disrupts temporal homeostasis but also contributes to pathological processes, including carcinogenesis, by affecting genes involved in cell proliferation, apoptosis, and DNA repair [67,68,69,70,71].

3.1. DNA Methylation and Histone Modifications

DNA methylation and histone modifications are key epigenetic mechanisms that regulate gene expression by altering chromatin accessibility and transcriptional activity [72,73]. DNA methylation is the addition of a methyl group to the fifth carbon of cytosine residues within CpG dinucleotides, catalyzed by DNA methyltransferases (DNMT1, DNMT3A, and DNMT3B), this modification commonly occurs in gene promoter regions and is generally linked to transcriptional repression [74]. In the context of circadian regulation, proper methylation patterns are essential for maintaining the rhythmic expression of core clock genes. However, exposure to PAHs has been shown to induce aberrant DNA methylation patterns, particularly hypermethylation of promoter regions of circadian genes such as BMAL1 and PER2, leading to their transcriptional silencing [26,75,76]

In addition to DNA methylation, histone modifications including acetylation, methylation, phosphorylation, and ubiquitination, play a critical role in regulating chromatin structure and gene expression [77,78]. Histone acetylation, typically associated with transcriptional activation, is dynamically regulated by histone acetyltransferases (HATs) and histone deacetylases (HDACs), many of which are themselves under circadian control. PAH exposure has been reported to disrupt the balance between these enzymes, leading to altered histone modification patterns at circadian gene loci [26,28]. For instance, reduced histone acetylation at CLOCK and BMAL1 promoters can limit transcriptional activation, while aberrant histone methylation may further reinforce gene silencing. These chromatin remodeling events contribute to long-term disruption of circadian oscillations and may persist even after cessation of exposure [67,79,80,81].

3.2. Non-Coding RNA Regulation of Circadian Genes

Non-coding RNAs, particularly microRNAs (miRNAs), represent another important layer of post-transcriptional regulation influencing circadian gene expression [82]. miRNAs are short non-coding RNA sequences (~22 nucleotides), endogenous RNA molecules that bind to complementary sequences in target messenger RNAs (mRNAs), leading to mRNA degradation or translational repression. Several miRNAs have been identified as key regulators of circadian clock genes, forming intricate regulatory networks that fine-tune circadian rhythms [83,84].

Exposure to PAHs has been shown to alter the expression profiles of numerous miRNAs, thereby affecting circadian gene regulation. For example, certain PAH-induced miRNAs can target transcripts of PER, CRY, and BMAL1, reducing their expression and disrupting the balance of the circadian feedback loop [85]. Additionally, miRNAs such as miR-34a, miR-132, and miR-155 have been implicated in both circadian regulation and inflammatory signaling, suggesting a convergence of pathways through which PAHs exert their effects [68,86]. Beyond miRNAs, long non-coding RNAs (lncRNAs) also contribute to circadian regulation by interacting with chromatin-modifying complexes and transcription factors, further influencing gene expression patterns [87,88,89]. The dysregulation of these non-coding RNAs by PAHs adds another layer of complexity to circadian disruption and highlights the importance of post-transcriptional mechanisms in environmental chronotoxicology.

3.3. Transcriptional Reprogramming of Core Clock Genes

The combined effects of epigenetic alterations and non-coding RNA regulation culminate in the transcriptional reprogramming of core circadian clock genes [66,90]. Central components of the circadian machinery, including CLOCK, BMAL1, PER (PER1–3), and CRY (CRY1–2), exhibit altered expression patterns following PAH exposure. These changes may manifest as either upregulation or downregulation, depending on the context, duration of exposure, and cellular environment [5,26,28].

The CLOCK gene, which encodes a transcription factor essential for initiating circadian gene expression, may be functionally impaired through reduced transcriptional activity or altered protein stability [91,92]. Similarly, BMAL1, a critical partner of CLOCK, is frequently downregulated in response to epigenetic silencing or transcriptional repression, leading to diminished activation of downstream target genes [93]. The PER and CRY genes, which form the negative feedback arm of the circadian loop, are also susceptible to dysregulation [66,94]. Reduced expression of these genes can impair feedback inhibition, resulting in destabilization of circadian oscillations (Figure 5). Conversely, aberrant overexpression of certain clock genes may lead to asynchronous signaling and further disrupt rhythmicity. These transcriptional alterations collectively represent a reprogramming of the circadian system, shifting it from a tightly regulated oscillatory network to a dysregulated state [22].

3.4. Functional Consequences of Gene Upregulation and Downregulation

The epigenetic and transcriptional rewiring of circadian clock genes has significant functional consequences for cellular physiology. One of the most prominent outcomes is the loss of rhythmic gene expression, characterized by diminished or absent oscillations in clock-controlled genes. This loss of rhythmicity disrupts the temporal coordination of essential biological processes, including metabolism, DNA repair, and cell cycle progression. As a result, cells may experience increased susceptibility to damage and reduced capacity for recovery, creating a permissive environment for disease development [19,33,95,96,97].

Disruption of circadian homeostasis also affects cellular signaling pathways that regulate cell proliferation, apoptosis, metabolism DNA repair [98,99]. For instance, downregulation of PER and CRY genes has been associated with impaired DNA damage response and reduced apoptosis, allowing damaged cells to survive and proliferate. Similarly, altered expression of BMAL1 can influence metabolic pathways and oxidative stress responses, further contributing to cellular dysfunction. The cumulative effect of these alterations is a breakdown of cellular homeostasis, which not only facilitates tumor initiation but also promotes tumor progression and resistance to therapy. Importantly, these changes are often sustained through epigenetic memory, highlighting the long-term impact of environmental exposures on circadian regulation and disease risk (Figure 6) [67,68,100,101,102,103].

Table 4. Epigenetic Modifications of Circadian Clock Genes Induced by PAHs [68,105,106,107,108,109].

Epigenetic Mechanism	Target Genes	Types of Modification	Effect on Expression	Functional Outcome
Histone Modification	CLOCK, BMAL1	Reduced acetylation, altered methylation	Decreased transcription	Impaired feedback loops
miRNA Regulation	PER, CRY, BMAL1	miRNA-mediated repression	mRNA degradation/inhibition	Disrupted oscillatory balance
lncRNA Interaction	Multiple clock genes	Chromatin remodeling	Variable expression changes	Long-term transcriptional reprogramming
Epigenetic Memory	Global circadian network	Persistent modifications	Sustained dysregulation	Increased disease susceptibility

Table 4. Epigenetic Modifications of Circadian Clock Genes Induced by PAHs [68,105,106,107,108,109].

Epigenetic Mechanism	Target Genes	Types of Modification	Effect on Expression	Functional Outcome
Histone Modification	CLOCK, BMAL1	Reduced acetylation, altered methylation	Decreased transcription	Impaired feedback loops
miRNA Regulation	PER, CRY, BMAL1	miRNA-mediated repression	mRNA degradation/inhibition	Disrupted oscillatory balance
lncRNA Interaction	Multiple clock genes	Chromatin remodeling	Variable expression changes	Long-term transcriptional reprogramming
Epigenetic Memory	Global circadian network	Persistent modifications	Sustained dysregulation	Increased disease susceptibility

4. Implications for Lung and Breast Cancer Initiation and Progression

The intersection between polycyclic aromatic hydrocarbon (PAH) exposure and circadian disruption has profound implications for cancer biology, particularly in tissues such as the lung and breast that exhibit heightened susceptibility to environmental and hormonal perturbations [26]. The ability of PAHs to induce DNA damage, alter epigenetic landscapes, and disrupt circadian regulation creates a multifactorial framework that promotes tumor initiation and progression. Importantly, circadian clock genes are not merely passive timekeepers but active regulators of cellular processes that maintain genomic integrity and tissue homeostasis. Their dysregulation therefore represents a critical mechanistic link between environmental toxicants and carcinogenesis [5,101].

4.1. Tissue-Specific Susceptibility to PAHs

The lung represents one of the primary targets of PAH toxicity due to its direct exposure to airborne pollutants [110]. Inhalation of PAH-containing particulate matter allows these compounds to deposit in the respiratory epithelium, where they undergo metabolic activation to form reactive intermediates capable of inducing DNA adducts and mutations. The high expression of cytochrome P450 enzymes such as CYP1A1 in lung tissue further enhances the bioactivation of PAHs, increasing the risk of carcinogenic transformation. Additionally, circadian regulation of pulmonary function and xenobiotic metabolism suggests that disruption of clock genes in lung cells may exacerbate susceptibility to PAH-induced damage, thereby promoting lung carcinogenesis (Figure 7) [9,101].

In contrast, the breast exhibits susceptibility to PAHs through mechanisms linked to endocrine and metabolic regulation [111]. Breast tissue is highly responsive to hormonal signaling, particularly estrogen, which itself is subject to circadian control. Estrogen receptors (ERα) are present in normal breast epithelial cells and exhibit variable activity, with their expression regulated in a circadian manner by molecular clock components, including the induction of PER2 [112,113,114,115]. PAHs have been shown to possess endocrine-disrupting properties, capable of interfering with estrogen receptor (ER) signaling and altering hormone metabolism [116,117]. Furthermore, lipophilic PAHs can accumulate in adipose-rich breast tissue, prolonging exposure and increasing the likelihood of cellular damage. Circadian disruption in breast tissue can impair hormonal regulation and metabolic homeostasis, creating conditions that favor tumor initiation and progression (Figure 8). This dual influence of environmental exposure and hormonal sensitivity underscores the vulnerability of breast tissue to PAH-induced carcinogenesis [38,107,118,119].

4.2. Circadian Regulation of Cell Cycle, DNA Repair, and Apoptosis

Circadian clock genes play a fundamental role in regulating key cellular processes that safeguard genomic integrity, including cell cycle progression, DNA repair, and apoptosis. These processes are tightly coordinated in a time-dependent manner to ensure that DNA replication and repair occur under optimal conditions [21,101,120]. To date, at least nine core circadian clock genes have been identified, including PER1, PER2, PER3, CLOCK, CRY1, CRY2, BMAL1, CK1ε, and TIM [121]. Core circadian regulators such as BMAL1, CLOCK, PER, and CRY influence the expression of genes involved in cell cycle checkpoints, including p53, p21, and cyclins, thereby controlling the timing of cell division [5].

Disruption of circadian regulation by PAHs can impair these protective mechanisms. For instance, downregulation of PER and CRY genes has been associated with defective DNA damage response pathways, leading to inefficient repair of DNA lesions. Similarly, alterations in BMAL1 expression can affect the transcriptional regulation of tumor suppressor genes, compromising their ability to control cell proliferation. The circadian clock also modulates apoptotic pathways, ensuring the elimination of damaged cells (Table 5). However, its disruption may result in reduced apoptosis and increased survival of genetically compromised cells. Collectively, these alterations contribute to genomic instability, a hallmark of cancer development [33,101,122,123].

4.3. Mechanisms of Tumor Initiation

Tumor initiation in the context of PAH exposure and circadian disruption is driven by the accumulation of genetic and epigenetic alterations. The metabolic activation of PAHs generates reactive intermediates that form DNA adducts, leading to mutations if not properly repaired [128,129]. In the presence of circadian disruption, the efficiency of DNA repair mechanisms is compromised, allowing these mutations to persist and accumulate over time. This process is further exacerbated by oxidative stress, which introduces additional DNA damage and promotes mutagenesis [14,28,130].

Epigenetic silencing of protective genes represents another critical mechanism in tumor initiation. Aberrant DNA methylation and histone modifications induced by PAHs can suppress the expression of tumor suppressor genes and circadian regulators such as PER2 and BMAL1 [131,132,133]. The loss of these protective functions disrupts normal cellular control mechanisms, facilitating uncontrolled cell proliferation and the early stages of tumorigenesis. Importantly, these epigenetic changes may be stable and heritable, contributing to long-term cancer risk even after exposure has ceased [67].

4.4. Tumor Progression and Metastasis

Beyond initiation, circadian disruption plays a significant role in tumor progression and metastasis. Dysregulation of circadian genes can enhance cellular proliferation by altering the expression of growth-promoting pathways, including those involving MYC and AKT signaling. Reduced expression of PER and CRY genes has been linked to increased tumor cell proliferation and resistance to apoptosis, enabling tumor expansion [5,33,104,134,135].

Circadian disruption also influences the tumor microenvironment, which plays a crucial role in cancer progression. Alterations in circadian regulation can affect immune responses, angiogenesis, and metabolic pathways within the tumor microenvironment [136]. Dysregulated circadian signaling may impair immune surveillance, allowing tumor cells to evade detection and destruction. Additionally, changes in metabolic rhythms can support the increased energy demands of rapidly proliferating tumor cells. These factors collectively contribute to enhanced invasion and metastasis, particularly in aggressive cancers such as lung and breast cancer [38,81].

While PER and CRY genes are predominantly downregulated, contributing to loss of circadian control and tumor suppression, components such as CLOCK and certain nuclear receptors (REV-ERB and ROR) may exhibit context-dependent upregulation, thereby facilitating tumor-promoting transcriptional and metabolic reprogramming [137,138,139,140].

4.5. Circadian Genes as Biomarkers in Cancer

Circadian clock genes are increasingly being recognized as valuable biomarkers for cancer diagnosis, prognosis, and therapeutic response [142]. Altered expression patterns of genes such as BMAL1, PER2, and CRY1 have been observed in both lung and breast cancers, with correlations to tumor stage, aggressiveness, and patient survival. Disruption or reduced expression of PER2 has been associated with poor prognosis in breast cancer (BC), while genetic loss or dysregulation of BMAL1 and per2 has been linked to tumor progression in lung cancer [122,126,143].

The potential of circadian genes as biomarkers extends to their use in precision medicine. Understanding the circadian profile of tumors may enable the development of time-based therapeutic strategies, optimizing the timing of drug administration to maximize efficacy and minimize toxicity [144,145]. Additionally, targeting circadian pathways may offer novel therapeutic approaches for cancer treatment. As research in this area continues to evolve, integrating circadian biology into clinical practice holds promise for improving cancer diagnosis and treatment outcomes [101,135].

A growing body of experimental and epidemiological evidence supports the mechanistic link between PAH exposure, circadian gene dysregulation, and cancer development. As summarized in Table 6, studies across cellular, animal, and human systems consistently demonstrate that disruption of core clock genes such as PER2, BMAL1, and CRY contributes to tumor initiation and progression, particularly in lung and breast tissues.

5. Therapeutic Implications and Future Directions

The growing recognition that circadian disruption is a central mediator of environmental carcinogenesis has opened new avenues for therapeutic intervention. In the context of polycyclic aromatic hydrocarbon (PAH)-induced toxicity, where molecular crosstalk between xenobiotic signaling, epigenetic remodeling, and circadian dysregulation drives disease progression, targeting temporal biology offers a promising strategy for improving cancer prevention and treatment. The integration of chronobiology into oncology has given rise to innovative approaches such as chronotherapy, circadian-based drug targeting, and epigenetic modulation, all of which aim to restore physiological rhythmicity and enhance therapeutic efficacy [101,145,179,180].

5.1. Chronotherapy and Circadian-Based Treatment Strategies

Chronotherapy refers to the strategic timing of drug administration in alignment with the body’s circadian rhythms to maximize therapeutic efficacy while minimizing toxicity [181]. This approach is based on the understanding that pharmacokinetics and pharmacodynamics are influenced by circadian oscillations in metabolism, hormone levels, and cellular sensitivity [182,183]. In cancer therapy, the timing of chemotherapeutic agents can significantly affect treatment outcomes, as tumor cells and healthy tissues often exhibit distinct circadian patterns of proliferation and repair [184].

In the context of circadian disruption induced by PAHs, chronotherapy holds particular relevance. Restoring or aligning treatment schedules with residual circadian rhythms may help counteract the deleterious effects of chronodisruption [26,28,185]. Furthermore, targeting circadian pathways directly represents an emerging therapeutic strategy. Pharmacological modulators of core clock components, including agonists or antagonists of REV-ERBα (NR1D1), REV-ERBβ (NR1D2) and ROR nuclear receptors, have shown potential in preclinical models for regulating metabolic and inflammatory pathways associated with cancer progression. By modulating circadian regulators, these approaches aim to re-establish rhythmic gene expression and suppress tumor-promoting processes [179,186].

5.2. Epigenetic Therapeutics and Clock Gene Modulation

Given the prominent role of epigenetic alterations in PAH-induced circadian disruption, epigenetic therapeutics offer a compelling avenue for intervention. DNA methylation inhibitors, such as DNA methyltransferases (DNMTs), including 5-azacytidine (marketed as Vidaza) and 5-Aza-2′-deoxycytidine (decitabine), have been widely studied for their ability to reverse aberrant promoter hypermethylation and restore the expression of silenced tumor suppressor genes such as (TP53) (p53), (BRCA1/2), (RB1) (Rb), and (PTEN), and circadian regulators such as PER2 and BMAL1. By reactivating these genes, DNA methylation inhibitors may help re-establish normal circadian function and inhibit tumor growth [93,187,188,189,190].

Similarly, drugs targeting histone modifications, including histone deacetylase (HDAC) inhibitors and histone methyltransferase inhibitors, can modulate chromatin structure and gene expression. HDAC inhibitors, in particular, have been shown to enhance histone acetylation at circadian gene promoters, thereby promoting transcriptional activation and restoring rhythmicity. These agents may also exert anticancer effects by inducing cell cycle arrest and apoptosis [191,192]. Importantly, the interplay between epigenetic regulation and circadian control suggests that combining epigenetic therapies with chronotherapy could provide synergistic benefits, enhancing treatment efficacy while reducing adverse effects [67,80,81,193].

5.3. Emerging Biomarkers and Clinical Translation

Circadian clock genes and their downstream targets are increasingly being explored as biomarkers for cancer diagnosis, prognosis, and therapeutic response. Altered expression patterns of genes such as BMAL1, PER2, and CRY1 have been associated with tumor progression and patient outcomes in lung and breast cancers. The integration of circadian biomarkers into clinical practice could improve early detection strategies by identifying individuals at high risk due to environmental exposures and circadian disruption [122,126,143].

Advances in high-throughput technologies have facilitated the identification of circadian signatures at the genomic, transcriptomic, and epigenomic levels. These signatures can be leveraged to develop personalized medicine approaches, in which treatment strategies are tailored based on an individual’s circadian profile and molecular characteristics. Determining the optimal timing of drug administration based on a patient’s circadian rhythm may enhance therapeutic outcomes. Additionally, monitoring circadian gene expression could provide insights into treatment response and disease progression, enabling more precise and adaptive clinical management [101,185].

5.4. Knowledge Gaps and Future Research Directions

Despite significant advances in understanding the relationship between PAH exposure, circadian disruption, and cancer, several knowledge gaps remain. One major limitation is the lack of longitudinal studies that track the temporal dynamics of circadian disruption and its long-term impact on disease development [194]. Most existing studies rely on cross-sectional or short-term experimental models, which may not fully capture the chronic effects of environmental exposures.

Mechanistic studies are also needed to further elucidate the complex interactions between AhR signaling, epigenetic regulation, and circadian clock machinery. In particular, the integration of multi-omics approaches including genomics, epigenomics, transcriptomics, proteomics, and metabolomics will be essential for providing a comprehensive understanding of these interactions. Systems biology frameworks can help model the dynamic networks underlying circadian regulation and identify key nodes for therapeutic targeting [6,195].

Additionally, there is a need to investigate the influence of exposure timing on PAH toxicity, as the concept of chronotoxicity suggests that the biological impact of toxicants may vary depending on the time of exposure. Understanding these temporal aspects could inform both risk assessment and intervention strategies. Finally, translating findings from experimental models to human populations remains a critical challenge, highlighting the importance of interdisciplinary research that bridges basic science, clinical studies, and public health.

5.5. Conclusion

The interplay between PAHs exposure and circadian disruption represents a critical axis in the pathogenesis of lung and breast cancer. PAHs exert their carcinogenic effects not only through direct genotoxic mechanisms but also by reprogramming circadian clock systems via AhR signaling, oxidative stress, inflammatory pathways, and epigenetic modifications. This disruption leads to the loss of rhythmic gene expression, impaired DNA repair, and dysregulation of cell cycle and apoptotic pathways, thereby facilitating tumor initiation and progression in lung and breast respectively.

The integration of circadian biology into environmental toxicology provides a more comprehensive framework for understanding cancer development and highlights novel opportunities for intervention. Therapeutic strategies such as chronotherapy, circadian pathway targeting, and epigenetic modulation offer promising avenues for improving cancer treatment outcomes. Furthermore, the identification of circadian biomarkers and the application of personalized medicine approaches have the potential to enhance early detection and optimize therapeutic strategies.

Ultimately, advancing our understanding of environmental chronotoxicology will require interdisciplinary efforts that integrate molecular biology, toxicology, chronobiology, and clinical research. By bridging these fields, it may be possible to develop more effective strategies for preventing and treating cancers (especially in lung and breast) associated with environmental exposures, thereby improving public health outcomes and advancing precision oncology (Figure 9).